Cartilage is composed of chondrocytes and a large quantity of intercellular cartilage matrices. Cartilage is classified into three types based on differences in cartilage matrix properties: i.e., hyaline cartilage constituting skeletal primordias at the fetal stage or major parts of articular cartilage; fibrocartilage comprising type I collagen in the matrix; and elastic cartilage existing in the auricula and in the epiglottis (see Shih Hone no Kagaku (Bone Biology), written and edited by Tateo Suda et al., Ishiyaku Pub, Inc. and KouSoshiki Kenkyu Handbook (Handbook of Hard Tissue Research), Division of Hard Tissue Research, Graduate School, Matsumoto Dental University).
Chondrocytes as constituents originate from pluripotent undifferentiated mesenchymal stem cells, and undifferentiated mesenchymal stem cells are aggregated by the Sox transcription factor (SRY-related HMG box-containing gene) 9 and converted into prechondrocytes. The prechondrocytes secrete a group of matrices such as type II collagen, type IX collagen, and proteoglycan to be converted into cartilage matrices. Consequently cartilage matrices increase, cells are individualized, and differentiated into chondroblasts through synergistic action with transcription factors such as Sox5 and Sox6. Further, transcription factor expression (i.e., of the runt-related gene 2 (Runx2)) is suppressed and expression of ERG/C-1-1 (i.e., the ets related gene) is accelerated in immature chondroblasts, and differentiation into permanent chondrocytes advances (see Hone/nankotsu taisha to chumoku no hone shikkan (Bone/cartilage metabolism and critical bone diseases), Toshio Matsumoto (ed.), Yodosha Co., Ltd. and Inada et al., Dev. Dyn 214: 279, 1999). It was reported that enlarged chondrocytes would lead to enhanced expression of type X collagen.
There are two patterns for the processes of cartilage growth. One of them is interstitial growth, in which cells differentiated, into chondrocytes and surrounded by the cartilage matrix proliferate through cell division. Each chondrocyte secretes a matrix, and cartilage tissue is then enlarged.
The other growth pattern is appositional growth caused by the perichondrium. Cartilage tissue is covered with a perichondrium except for the articular surface of the articular cartilage. The strong perichondrium is constituted of fibroblasts, but is similar to chondrocytes in the inner layer, thus the difference between fibroblasts and chondrocytes is unclear. Perichondrium cells in the inner layer proliferate while gradually changing into circular forms, and such cells further a secrete cartilage matrix and grow outwardly.
In general, the outside of the perichondrium of the cartilage tissue is in contact with connective tissue that is rich in blood vessels and nerves, although the inside thereof does not contain blood vessels or nerves. When the cartilage is damaged inside, accordingly, it cannot be repaired by undifferentiated stem cells, cytokines, and so on. In addition, the capacity of the chondrocytes for mitotic proliferation is poor, and self-repair of chondrocytes is very difficult.
The method to repair a articular cartilage is a topical injection in cartilage matrix, a implantation of cultured bone marrow mesenchymal stem cells to prepare autologous chondrocytes, or an impregnating of matrix disc with chondrocyte growth factors. Transforming growth factor (TGF-β1), insulin-like growth factor (IGF-1), basic fibroblast growth factor (bFGF), PTH-related peptide (PTHrP) that are highly homologous to the 13 N-terminal amino acids of PTH, hepatocyte growth factor (HGF), and bone morphogenetic protein (BMP) belonging to TGF-β superfamily have been reported as chondrocyte growth factors. BMP and TGF-β may induce to lose the function of chondrocytes after implantation due to calcification in article cartilage, because these factors have some important effects, such as aggregation of undifferentiated mesenchymal stem cells, and inhibitory effect on terminal chondrocytic differentiation in addition to the effect on chondrocyte growth.
Osteoclast differentiation factor (i.e., the receptor activator of NF-κB ligand (RANKL)) is a membrane-binding protein of tumor necrosis factor (TNF) family that is induced on osteoblasts/stromal cells by bone resorption factors, and is necessary for differentiation and maturation of osteoclasts (see Yasuda et al., Proc. Natl. Acad. Sci., U.S.A., 95: 3597, 1998 and Lacey et al., Cell 93: 165, 1998). Research focusing on RANKL/RANK/OPG, including a receptor (i.e., the receptor activator of NF-κB (RANK)) and a decoy receptor (i.e., the osteoprotegerin (OPG)), has led to elucidation of the control mechanism for osteoclast differentiation and maturation in vivo, and the correlation between these 3 molecules and metabolic bone diseases has also become elucidated (see Suda et al., Endocr Rev, 20: 345, 1999).
A correlation between RANKL and differentiation and proliferation of prechondrocytes and/or mesenchymal stem cells has been unknown.